Synthetic cornea from retinal stem cells
First Claim
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1. An isolated retinal-stem cell derived synthetic cornea.
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Abstract
Methods of producing synthetic corneas are disclosed which are differentiated from retinal stem cells (rSC) derived from parthenogenetically activated human oocytes, including that such synthetic corneas are produced in the absence of a 3-D scaffold. Isolated synthetic corneas, produced by the disclosed methods, are also described.
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Citations
36 Claims
- 1. An isolated retinal-stem cell derived synthetic cornea.
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13. A method of producing a synthetic cornea comprising:
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a) parthenogenetically activating a human oocyte, wherein activating comprises;
i) contacting the oocyte with an ionophore at high O2 tension and ii) contacting the oocyte with a serine-threonine kinase inhibitor at low O2 tension;b) cultivating the activated oocyte of step (a) at low O2 tension until blastocyst formation; c) transferring the blastocyst to a layer of feeder cells, and culturing the transferred balstocyst under high O2 tension; d) mechanically isolating an inner cell mass (ICM) from trophectoderm of the blastocyst of step (c); e) culturing the cells of the ICM of step (d) on a layer of human feeder cells under high O2 tension, wherein retinal stem cells are identified in the culture by human embryonic stem cell markers (hES) and neuron specific markers, and wherein the identified retinal stem cells are subsequently isolated; f) culturing the isolated stem cells of step (e) in media comprising serum replacement (M/SR), plasmonate, and at least one mitogen that activates the gp130/STAT pathway and/or the MAP kinase pathway on a fibroblast feeder layer treated with a DNA synthesis inhibitor; g) culturing the mitogen treated cells of step (f) in M/SR comprising plasmonate (M/SRP), without added mitogen, to near confluence, wherein ½
volume of the M/SRP is replaced with M/SR periodically until the near confluent cells develop pigmentation and a domed appearance; andh) transferring the pigmented cells of step (g) in M/SR to a gelatin coated substrate, wherein ½
volume of the M/SR is replaced with M/SR periodically until a synthetic cornea develops. - View Dependent Claims (14, 15, 16, 17, 18, 19, 20, 21)
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- 22. A method of treating a subject in need thereof, comprising replacing a cornea of the subject with a synthetic cornea.
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26. A method of identifying an agent that affects the cornea of an eye comprising:
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(a) contacting a retinal-stem cell derived synthetic cornea with an agent; and (b) observing a change to the cornea in the presence and absence of the agent, wherein a change to the cornea is indicative of an agent that affects the cornea. - View Dependent Claims (27, 28, 29, 30, 31, 32)
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Specification